Infection of animal cells by herpes simplex virus (HSV) can lead to a productive lytic infection in which viral replication and gene expression are tightly coupled processes. A better understanding of the interaction between these processes has direct relevance to our understanding of the molecular mechanisms between regulate cell cycle control. Therefore, a major DNA binding protein of HSV, infected cell (ICP8), will be studied for its role in the transition between these processes. ICP8 is essential for viral DNA replication and plays a major regulatory role in HSV gene expression. Furthermore, the ability of ICP8 to down-regulate or to stimulate gene expression from viral late promoters is dependent on the replication status of the viral genome. To examine this paradox, ICP8's role in the stimulation of late gene expression will be investigated in several ways. These approaches include examination of the structure of viral chromatin in ICP8 mutants and determination of interactions between ICP8 and host cellular proteins. Techniques to analyze viral gene expression and protein-protein interactions will include in vivo RNA labeling, Northern hybridizations, the yeast two- hybrid system and/or GST fusions. These studies will yield both basic information about the mechanisms of gene regulation during HSV infection, as well as provide important information regarding the regulation of cell cycle control.